Journal: bioRxiv

Article Title: Chondrocyte-specific RUNX2 Overexpression Causes Chondrodysplasia During Development, but is Not Sufficient to Induce OA-like Articular Cartilage Degeneration in Adult Mice Without Injury

doi: 10.1101/470005

Figure Lengend Snippet: Chondrocyte-specific RUNX2 overexpression during development leads to chondrodysplasia. (A) Alcian blue/Alizarin red skeletal staining of E18.5 R26 Runx2/+ (Control), Col2a1 CreERT2/+ ; R26 Runx2/+ ( C2T2; R26 Runx2/+ ), and Col2a1 CreERT2/+ ; R26 Runx2/Runx2 ( C2T2; R26 Runx2/Runx2 ) embryos. (B) Femur (left), tibia (middle), and humerus (right) lengths from E18.5 Control, C2T2; R26 Runx2/+ , and C2T2; R26 Runx2/Runx2 embryos (n = 3 to 5 for all groups). * p < 0.05, ** p < 0.01, *** p < 0.001, one-way ANOVA followed by Dunnett’s multiple comparisons test. (C) Alcian blue Hematoxylin/Orange G staining of humerus growth plate sections from E18.5 Control, C2T2; R26 Runx2/+ , and C2T2; R26 Runx2/Runx2 embryos. Top panels are 5X images of the proximal humerus; middle panels are high magnification images (20X) of the proliferating and pre-hypertrophic zones (yellow boxes); bottom panels are high magnification images (20X) of the hypertrophic zone (orange boxes). (D) In situ hybridization for Col10a1 (top panels) and Mmp13 (bottom panels) on E18.5 Control, C2T2; R26 Runx2/+ , and C2T2; R26 Runx2/Runx2 proximal humerus sections. (E) TUNEL staining of proximal humerus sections from E18.5 Control, C2T2; R26 Runx2/+ , and C2T2; R26 Runx2/Runx2 embryos (5X images). (B) Quantification of TUNEL + cell number as percentage of total cell number within the growth plate of the proximal humeri of E18.5 Control, C2T2; R26 Runx2/+ , and C2T2; R26 Runx2/Runx2 embryos (n = 3 for all groups). * p < 0.05, one-way ANOVA followed by Dunnett’s multiple comparisons test.

Article Snippet: Sections were rinsed in phosphate buffered saline + 0.1% tween-20 (PBST) and incubated with ImmPress HRP anti-rabbit IgG peroxidase polymer secondary (anti-MMP13, Vector Labs) or ImmPress HRP anti-mouse IgG peroxidase polymer secondary (anti-COL10A1, Vector Labs) for 30 minutes.

Techniques: Over Expression, Staining, In Situ Hybridization, TUNEL Assay

Journal: bioRxiv

Article Title: Chondrocyte-specific RUNX2 Overexpression Causes Chondrodysplasia During Development, but is Not Sufficient to Induce OA-like Articular Cartilage Degeneration in Adult Mice Without Injury

doi: 10.1101/470005

Figure Lengend Snippet: Postnatal RUNX2 overexpression induces MMP13 expression in unmineralized articular chondrocytes prior to accelerating articular cartilage degeneration following MLI. (A) Safranin O/Fast green staining of knee joint sections from R26 Runx2/+ (Control) and Acan CreERT2/+ ; R26 Runx2/+ (RUNX2 OE) mice injected with tamoxifen at 2 months of age and subjected to sham or MLI at 2.5 months of age. Joints were harvested one month following injury. Top panels are 5X images of the knee joint; bottom panels are high magnification images (20X) of the boxed regions. (B) Cartilage degeneration in Control (n = 6) and RUNX2 OE (n = 6) mice was evaluated by modified OARSI scoring (top graph) and quantitative histomorphometric analysis (bottom graph, data normalized to contralateral sham control sample). (C) MMP13 (left panels) and COL10A1 (right panels) immunohistochemistry of knee joint sections from Control or RUNX2 OE mice subjected to sham or MLI at 2.5 months of age and harvested 1 month following injury (20X images). (D) Quantification of MMP13 + cell number as percentage of total cell number from Control and RUNX2 OE mice (MLI samples only, n = 4 for both groups). (E) Quantification of COL10A1 + cartilage area as percentage of total cartilage area from Control and RUNX2 OE mice (MLI samples only, n = 4 for both groups). * p < 0.05, Student’s t-test.

Article Snippet: Sections were rinsed in phosphate buffered saline + 0.1% tween-20 (PBST) and incubated with ImmPress HRP anti-rabbit IgG peroxidase polymer secondary (anti-MMP13, Vector Labs) or ImmPress HRP anti-mouse IgG peroxidase polymer secondary (anti-COL10A1, Vector Labs) for 30 minutes.

Techniques: Over Expression, Expressing, Staining, Injection, Modification, Immunohistochemistry